The HMG-I/Ygene encodes the HMG-I and -Y protein isoforms, which function as architectural chromatin binding proteins involved in transcriptional regulation. These proteins are up-regulated in human cancer, although their role in the pathogenesis of malignancy is unclear. To understand how HMG-I/Y proteins may contribute to transformation, we are exploring their regulation and function. We discovered that HMG-I/Y is a direct c-Myc target gene important in Burkitt's lymphoma. We also demonstrated that HMG-I/Y is necessary for transformation because decreasing these proteins in human cancer cell lines blocks the transformed phenotype. We were the first to show that HMG-I/Y proteins have several oncogenic properties. Specifically, overexprossion of HMG-I or-Y leads to anchorage-independent cell growth in several experimental cell lines. Fibroblasts overexpressing HMG-I or-Y are tumorigenic in nude mice. We developed transgenic mice overexpressing HMG-/in lymphoid cells and all of them develop lymphoid hyperplasia and malignancy at a mean age of 8 months. HMG-I overexpression also correlates with genomic instability, which may contribute to tumor initiation or progression. Thus, I hypothesize that HMG-I/Y is an oncogene important in the pathogenesis of human cancer. The focus of this research proposal is to identify the mechanisms involved in transformation by HMG-I/Y using unique reagents developed in my laboratory. Our Specific Aims Are: 1.) Identify and characterize direct HMG-I/Y gene targets involved in neoplastic transformation using microarray analysis. 2.) Define the functional domains of HMG-I/Y involved in transformation, chromosomal instability, and cell cycle regulation. A.) Identify the functional domains of HMG-I/Y required for transformation using the soft agar transformation assay. B.) Investigate the role of HMGI/ Y in genomic instability and identify the relevant domains using spectral karyotyping analysis. C.) Investigate the role of HMG-I/Y in cell cycle regulation and identify the relevant domains using cell cycle profile analysis. 3.) Define the pathways involved in transformation using our HMG-I transgenic mice. A.) Assess transformed lymphoid cells from the HMG-I trangenic mice for overexpression of HMG-I target genes and chromosomal instability B.) Identify pathways involved in transformation by HMG-I by crossing the HMG-I transgenic mice with other genetically altered mice. 4.) Determine if HMG-I/Yexpression correlates with prognosis and clinical outcome in lymphoid malignancies and brain tumors. Study HMG-I/Y gene and protein expression in patient samples from the Johns Hopkins Leukemia and Brain Tumor Banks. This proposal is significant because the results will enhance our understanding of human malignancies with increased HMG-I/Y proteins and may lead to new treatment strategies.